The Epstein-Barr virus (EBV) infects B cells and establishes a latent infection that persists for the lifetime of the host. In vitro, infected B cells can proliferate indefinitely, resulting in the immortalization of these cells. Thus, EBV-mediated transformation is a prerequisite for any ensuing EBV-associated malignancy. We have demonstrated that one nuclear antigen of EBV, EBNA-3A, which is essential for transformation, interacts directly with the cellular DNA-binding factor, Jk. Additionally, we have shown that EBNA-3A binds a second cellular protein of unidentified function of approximately 80 kDa. This interaction appears to be specific for EBNA-3A because the corresponding region of EBNA-3C, which is also essential for transformation and binds to Jk, shows no interaction. Furthermore, this interaction does not seem to be restricted to a particular B cell line nor only to B cells. The objective of this project is to identify the role of EBNA-3A in EBV-mediated transformation. Specifically, we propose to identify this 80 kDa cellular protein using the yeast two-hybrid system and to evaluate the function and significance of its interaction with EBNA-3A, as it relates to EBV-mediated transformation. The significance of EBNA-3A within the context of viral infection will be addressed by generating recombinant EBV in which the known functions of EBNA-3A have been disrupted by specific mutations.